In vitro sensitivity of normal human mesothelial and malignant mesothelioma cell lines to four new chemotherapeutic agents

In this study, we used four human mesothelioma cell lines (M14K, M24K, M25K and M38K), one transformed human mesothelial cell line (MeT-5A) and one primary mesothelial culture (UPL) to test for in vitro sensitivity to docetaxel, paclitaxel, SN-38 [an active metabolite of irinotecan (CPT-11)] and gemcitabine, as single agents. Subconfluent cell cultures were treated with 2x10(-9), 5x10(-9), 10(-8), 2x10(-8) and 5x10(-8) M concentrations of each drug for 48 h. The sensitivity was measured in terms of cell viability using the Trypan blue exclusion method. All four drugs were potent inhibitors of mesothelioma cell growth, but cell lines from different patients diverged in their sensitivity to the individual agents. In most cases docetaxel, paclitaxel and SN-38 were more potent killers of mesothelioma cells than gemcitabine. The induction of DNA damage was investigated using the Comet assay; cells from two cell lines (M14K and M25K) were treated with subtoxic 10(-8) M concentrations of each drug for 4, 24 and 48 h. Each of the agents caused a slight increase in DNA single-strand breaks at a concentration of 10(-8) M.     

Enhanced efficiency of thermally targeted taxanes delivery in a human xenograft model of gastric cancer

Since successful chemotherapy with taxanes requires an improvement in their therapeutic index, especially by the reduction in unwanted systemic toxicity of either drug or adjuvants, we have investigated and are reporting results from an investigation of the use of a novel polymeric thermosensitive micellar delivery system for docetaxel and paclitaxel. Here we reported a novel metastable thermosensitive polymeric micelle for docetaxel and paclitaxel delivery [poly(N-isopropylacrylamide-co-acrylamide)-b-poly(DL-lactide), Poly(IPAAm-co-AAm)-b-PDLLA]. Previous in vitro efficacy studies indicated that, with hyperthermia, docetaxel-loaded micelles showed stronger cytotoxicity to different tumor cell lines than conventional docetaxel formulation while exhibiting slighter toxicity to normal cells. Present in vivo studies indicated that at the same dose level of docetaxel (paclitaxel), hyperthermia greatly enhanced the antitumor effect of micellar docetaxel (paclitaxel) in human gastric BGC mouse xenograft model by showing an extraordinary tumor volume and weight growth percentage inhibition of more than 80%. Meanwhile, acute toxicity tests features the lower LD(50) of the combination of hyperthermia and micellar docetaxel (paclitaxel) compared to that of the control group. The present results suggest that poly(IPAAm-co-AAm)-b-PDLLA micelles could be a clinically useful chemotherapeutic formulation and merit further research to evaluate the feasibility of clinical application.     

The clinical development of new mitotic inhibitors that stabilize the microtubule

Microtubule-stabilizing agents are increasingly studied for cancer treatment based largely on the prior success of paclitaxel and docetaxel. In this review, we focus on the clinical development of epothilones and discodermolide, and we discuss salient preclinical and clinical highlights of these two novel natural products. These agents are distinguished by their biochemical features making them poor P-glycoprotein substrates and capable of inducing cytotoxicity in cell lines or in vivo tumor models harboring mutations in tubulin. There is now considerable data regarding the efficacy of the epothilones in human beings and discodermolide holds such promise, as well.     

Signal transduction pathways of taxanes-induced apoptosis

Docetaxel (Taxotere) is a member of the taxane class of anticancer agents to reach clinical use. This semisynthetic analog of paclitaxel (Taxol) is one of the newer potent anti-neoplastic agents now undergoing extensive laboratory and clinical investigations. Several studies indicate that antimicrotubule agents are potent promoters of apoptosis in cancer cells. Cytotoxic mechanisms of antimitotic taxoids are not yet fully understood, but it has been demonstrated that docetaxel increases tubulin polymerisation, promotes microtubule assembly and also inhibits tubulin depolymerisation. Disruption of microtubules results also in the induction of tumor suppressor gene p53 and inhibitor of cyclin-dependent kinases and activation/inactivation of several protein kinases. As a consequence cells are arrested in the G2-M phase of the cell cycle, after which they may either undergo cell death by apoptosis or necrosis or overcome the G2-M stop and continue in the division cycle (often toward a post-mitotic cell death) depending on the tumor cell type. Nevertheless, how docetaxel induces apoptotic cell death or caspases activation is not yet defined. One may assume that taxanes are able to induce the phosphorylation of Bcl-X(L)/Bcl-2 members and thus inactivate their anti-apoptotic capacities. The down-regulation of Bcl-2 and/or the upregulation of p53 and p21/WAF-1 are certainly one of the important modes of apoptosis induction by taxanes. The aim of this framework is to summarize the effects of microtubuline targeting agents on apoptotic signal transduction and new molecular pathways. Finally, we will also discuss the potential therapeutic interest in the association of docetaxel and ionizing radiation.     

Some Anticancer Agents Act on Human Serum Paraoxonase‐1 to Reduce Its Activity

Human serum paraoxonase (hPON1) is an important antioxidant enzyme. It protects low‐density lipoproteins against oxidative stress and prevents atherosclerosis development. Anticancer agents have cardiotoxic effects, and this situation can lead to significant complications. Our aim was to evaluate the in vitro effects of some of the anticancer agents such as cetuximab, paclitaxel, etoposide, docetaxel, and ifosfamide on the activity of hPON1 in this study. For this reason, PON1 was purified from human serum with a specific activity of 3654.2 EU/mg and 16.84% yield using simple chromatographic methods. The five chemotherapeutic agents dose dependently decreased in vitro hPON1 activity. IC₅₀ values for cetuximab, paclitaxel, etoposide, docetaxel, and ifosfamide were 0.0111, 0.042, 0.226, 0.665, and 23.3 mm , respectively. K_i constants were 0.0194, 0.0165, 0.131, 0.291, and 8.973 mm , respectively. The inhibition mechanisms of cetuximab, etoposide, docetaxel, and ifosfamide were non‐competitive, and for paclitaxel was competitive. Consequently, inhibition of hPON1 by these anticancer agents may explain some of the cardiotoxic actions of these drugs.     

Determination of docetaxel and Paclitaxel in human plasma by high-performance liquid chromatography: validation and application to clinical pharmacokinetic studies

Taxanes, docetaxel and paclitaxel, represent important antineoplastic agents with broad spectra of antitumor activity. The authors developed and validated a high-performance liquid chromatography method with ultraviolet detection for quantifying both taxanes in human plasma. The assay uses liquid-liquid extraction as sample treatment and an isocratic mobile phase and reversed-phase chromatography to determine docetaxel with paclitaxel as internal standard and vice versa. The lower limit of quantification was 0.015 mg/L. The assay had good recovery (87.96+/-14.05 and 90.57+/-9.63 for docetaxel and paclitaxel respectively) and precision: the within-day and between-days relative standard deviation of the mean for docetaxel (0.015-3 mg/L) and paclitaxel was always <10%. The method presented has been fully validated following the U.S. Food and Drug Administration requirements and has been successfully applied for the pharmacokinetic investigation of docetaxel or paclitaxel.     

Structural studies of taxol analogues for drug discovery

Background: The major mechanism of action of antitumor taxanes, including two clinically useful antitumor agents, paclitaxel and docetaxel, lies in their interactions with β-tubulin in microtubule polymers, leading to cell cycle arrest and apoptosis. However, owing to the technical limitations, the molecular interactions of ligands with the residues in taxane binding sites of tubulin as well as the conformations adopted by taxanes on binding are still not fully understood. Objective: This review focuses on the exploration of paclitaxel's interactions with tubulin, and the impact of such efforts on the drug discovery for new taxanes and microtubule stabilizing agents (MSAs). Methods: Data were identified through the search of Chemical Abstracts and PubMed databases for research articles and reviews up to April 2008. Conclusion: Based on a collection of information gathered from crystallography, 1D and 2D NMR spectroscopy (NOESY, ROESY, REDOR), and structural–activity relationship (SAR) by chemical synthesis and pharmacological assays, ‘opened’ or T-shape conformations have been predicted to be the biologically active ones in recent years, and confirmed by further SAR studies. Some more potent analogues than paclitaxel or simplified compounds with similar potencies to that of paclitaxel have been discovered. Structural studies of taxol analogues will continue to make great contributions to the rational design of taxanes and novel prototype MSAs for drug discovery.     

Role of Formulation Vehicles in Taxane Pharmacology

The non-ionic surfactants Cremophor EL (CrEL) and Tween 80,both used as formulation vehicles of many (anticancer) agentsincluding paclitaxel and docetaxel, are not physiologicalinert compounds. We describe their biological properties,especially the toxic side effects, and their pharmacologicalproperties, such as modulation of P-glycoprotein activity. Indetail, we discuss their influence on the disposition of thesolubilized drugs, with focus on CrEL and paclitaxel, and ofconcomitantly administered drugs. The ability of thesurfactants to form micelles in aqueous solution as well asbiological fluids (e.g. plasma) appears to be of greatimportance with respect to the pharmacokinetic behavior of theformulated drugs. Due to drug entrapment in the micelles,plasma concentrations and clearance of free drug changesignificant leading to alteration in pharmacodynamiccharacteristics. We conclude with some perspectives related tofurther investigation and development of alternative methodsof administration.     

Docetaxel (taxotere) in the treatment of non-small cell lung cancer

Docetaxel is a new semi-synthetic anticancer agent derived from bacatin III of the needles of the European yew Taxus baccata. Docetaxel has a novel mechanism of action since it binds to tubulin inducing its polymerization and promoting stable microtubule formation. Several differences exist between docetaxel and paclitaxel: (i) broader activity of docetaxel against freshly explanted human tumors than paclitaxel; (ii) a 2-fold higher affinity than paclitaxel; (iii) 2.5-fold more potent than paclitaxel as an inhibitor of cell replication and (iv) docetaxel acts at the S-phase whereas paclitaxel at the G(2)/M phases of the cell cycle. Preclinical and phase II studies revealed that docetaxel is active against NSCLC. In chemotherapy-na ve patients with NSCLC response rates ranged from 19% to 54% with a median duration of survival ranging from 6.3 months to 11 months, and 1-year survival ranging from 21% to 71%. Docetaxel as single agent provided a survival as well as a clinical benefit over BSC in untreated patients with NSCLC. Docetaxel has been efficiently combined with cisplatin (ORR 33%-46%), carboplatin (ORR 30%-48%), vinorelbine (ORR 20%-51%), gemcitabine (ORR 37%-47%), with a median survival ranging from 5-14 months. A preliminary analysis of a multicenter randomized trial comparing docetaxel/CDDP with docetaxel/gemcitabine revealed that the two regimens had comparable activity in terms, of response rates, duration of response, TTP and overall survival; however, the docetaxel/gemcitabine combination has a most favourable toxicity profile compared to docetaxel/CDDP. Docetaxel has also demonstrated radiosensitizing properties and encouraging results have been achieved in combination with irradiation. Finally, docetaxel has shown an important activity in previously-treated patients with NSCLC with ORR ranging from 16% to 25% with a median survival ranging from 7.2 months to 10.5 months. Randomized trials revealed that second-line docetaxel confers a survival benefit over either BSC or ifosfamide/vinorelbine in pretreated patients with NSCLC.     

Effects of paclitaxel, docetaxel and their combinations on subcutaneous lymphomas in inbred Sprague–Dawley/Cub rats

We investigated, whether the effects on paclitaxel, docetaxel or their combinations on T-cell lymphomas in Sprague-Dawley/Cub rats were mainly caused by their different efficiency or combination of different mechanism of action, or limited by metabolic inactivation by P450 enzymes or drug efflux caused by P-glycoprotein (P-gp). Docetaxel most effectively prolonged the survival of rats and the time of lymphoma appearance, inhibited their intravital size and weight after sacrifice. Paclitaxel was poorly effective and combined administration had intermediate effects. Blood levels of both drugs were similar. Repeated administration of paclitaxel, but not docetaxel, decreased its area under concentration, but the effect disappeared 6h after dosing and was not sufficient to explain lower effects of paclitaxel. The faster metabolism of docetaxel than paclitaxel in vitro did not limit its higher efficiency and repeated administration of paclitaxel did not induce its metabolism to decrease its blood levels sufficiently. Likewise, undetectable expression of P-gp protein in tumours could not explain lower effects of paclitaxel, which is a better substrate of P-gp. Docetaxel was three-fold more effective than paclitaxel against P388D1 lymphoma cell line, used as a model of the T-cell lymphoma and combined action was dominated by the effects of docetaxel. Thus, docetaxel was effective against T-cell lymphomas and may be a potential anticancer drug in similar indications.     

Management of toxicities associated with the administration of taxanes

The taxanes (paclitaxel and docetaxel) are highly active cytotoxic antineoplastic agents. Common toxicities of the drugs include total alopecia, hypersensitivity reactions, bone marrow suppression (principally neutropenia), arthralgia, myalgias, and peripheral neuropathy. When administered as a 3-h infusion, paclitaxel appears to be associated with a lower risk of neutropenia and a greater risk of peripheral neuropathy, compared to either 24-h infusion paclitaxel or docetaxel (1-h infusion). Neither paclitaxel nor docetaxel is associated with a high risk for significant emesis. High cumulative doses of docetaxel have been shown to produce fluid retention (e.g., oedema, ascites, pleural effusions), while paclitaxel, when combined with doxorubicin, increases the risk of anthracycline-induced heart failure. Both paclitaxel and docetaxel have been administered at lower dose levels, on a weekly schedule, with acceptable toxicity profiles. In general, the side effects of the taxanes are manageable, and few patients discontinue treatment due to excessive toxicity.     

Management of toxicities associated with the administration of taxanes

The taxanes (paclitaxel and docetaxel) are highly active cytotoxic antineoplastic agents. Common toxicities of the drugs include total alopecia, hypersensitivity reactions, bone marrow suppression (principally neutropenia), arthralgia, myalgias, and peripheral neuropathy. When administered as a 3-h infusion, paclitaxel appears to be associated with a lower risk of neutropenia and a greater risk of peripheral neuropathy, compared to either 24-h infusion paclitaxel or docetaxel (1-h infusion). Neither paclitaxel nor docetaxel is associated with a high risk for significant emesis. High cumulative doses of docetaxel have been shown to produce fluid retention (e.g., oedema, ascites, pleural effusions), while paclitaxel, when combined with doxorubicin, increases the risk of anthracycline-induced heart failure. Both paclitaxel and docetaxel have been administered at lower dose levels, on a weekly schedule, with acceptable toxicity profiles. In general, the side effects of the taxanes are manageable, and few patients discontinue treatment due to excessive toxicity.     

LC-MS/MS法同时测定人血浆中紫杉醇和多西他赛的质量浓度

目的建立一种简单、快速,可同时测定人血浆中紫杉醇和多西他赛药物质量浓度的LC-MS/MS方法。方法以罗红霉素为内标,血浆经乙腈沉淀蛋白、离心后进样分析;色谱柱为Hypersil GOLD aQ;流动相为1mL·L-1甲酸溶液-乙腈;梯度洗脱;流速为0.4mL·min-1;离子源为可加热电喷雾离子源(HESI);检测方式:正离子检测;扫描方式为多反应监测(MRM),紫杉醇、多西他赛和罗红霉素的检测离子对分别为m/z854.3→509.1,808.3→527.1和837.6→679.3。结果每份含紫杉醇和多西他赛的样品分析时间为6min,血浆中紫杉醇和多西他赛的药物质量浓度在0.005~1.000μg·mL-1范围内线性关系良好,最低定量限为0.005μg·mL-1,两药测定的日内和日间精密度RSD<15%,提取回收率分别为94.97%~101.95%和96.07%~112.86%,基质效应分别为101.14%~115.59%和82.40%~95.77%。紫杉醇和多西他赛血浆样品在反复冻融3次、4℃保存2h和-80℃保存1个月的条件下稳定。结论该研究建立的LC-MS/MS分析方法简便、快速、灵敏和准确,能同时测定人血浆中紫杉醇和多西他赛的质量浓度。     

Current state of the art of new tubulin inhibitors in the clinic

For years the microtubule stabilizing agents docetaxel and paclitaxel belong to the most successful clinical chemotherapeutic agents. Several attempts have been made over the years to equal and better these drugs. Both taxanes are associated with the notorious side effect neurotoxicity and are often accompanied with increased drug resistance and cross resistance with other chemotherapeutic agents. In addition their high lipophilicity demands use of co-solvents, which are associated with less favorable side effects such as hypersensitivity. To prevent these disadvantages and improve the clinical application of the taxanes several new agents have entered clinical testing. The agents that are discussed are the drug class of the discodermolides; XAA296A and the epothilones; BMS-247550, BMS-310705, epo906, kos-862 and the agents ABT-751 and D-24851. Here we present an overview of recently performed clinical studies to determine the current state of the art of the tubulin inhibitors which are intended to enlarge and improve the clinical use of the taxanes docetaxel and paclitaxel.     

General and recent aspects of the chemistry and structure-activity relationships of taxoids

The antimitotic agents of the Taxoid series are important substances as anticancer drugs. The efficacy of paclitaxel (Taxol) and docetaxel (Taxotere) has been well demonstrated in the treatment of breast, ovarian and lung cancers. Although these drugs have brought benefits in cancer chemotherapy, they unfortunately possess some disadvantages due to their inefficacy on certain resistant cancers, and due to their toxic side effects. For these reasons, the synthesis of new taxoids with improved biological activity is still an important research area. This review while covering general aspects of taxoid chemistry focuses on recent developments in this area especially those contributing to the improved availability of antimitotic taxoids. Studies of structure-activity relationships are also described.     

Which dosing scheme is suitable for the taxanes? An in vitro model

The discovery and development of the taxane class of antitumor compounds represent significant advances in the treatment of patients with a variety of malignancies. These drugs are effectively used in the treatment of breast cancer. In this study we evaluated the efficacy of fractionated usage of both paclitaxel and docetaxel as a single agent in the breast cancer cell line MCF-7. It has been shown that the cytotoxic effect of paclitaxel was increased when the divided IC50 concentrations were used sequentially and in contrast to paclitaxel, cytotoxic effect of docetaxel was decreased with the same schema and the single dose of IC50 concentration was optimal. The cause of the difference between the cytotoxic effects of two agents with this schedule is obscure. Demonstrating mechanisms, which are responsible for these differences, will be important for more rational use of taxoids and to provide basis for the following clinical trials.     

Docetaxel. A pharmacoeconomic review of its use in the treatment of metastatic breast cancer.

Advanced breast cancer has a poor prognosis (median duration of survival about 2 years); thus, treatment options are largely palliative. Recent studies with the taxoids docetaxel and paclitaxel suggest that these agents are effective second-line therapy in patients with metastatic breast cancer. Objective response rates range from 30 to 57% with docetaxel monotherapy in clinical trials in patients with advanced disease who had failed to respond to previous therapy, and the median time to disease progression ranges from 17 to 20 weeks. Currently available pharmacoeconomic data relating to docetaxel in the treatment of women with metastatic breast cancer include a cost-minimisation study and 3 cost-utility studies which relate to the UK, French and US healthcare systems. All analyses compared docetaxel with paclitaxel. Since no direct clinical comparisons of these 2 agents have been performed, model assumptions were based on available, rather than comparative, clinical data. The French model also considered vinorelbine monotherapy as a comparator. Considering direct costs only, costs associated with docetaxel ranged from < 0.3% less to 13% more than those of paclitaxel. When utilities were evaluated, docetaxel was associated with incremental gains versus paclitaxel of 33 and 75 additional days of quality-adjusted health in the UK and US studies, respectively, and 22 additional days of quality-adjusted progression-free survival (QPFS) in the French analysis. Thus, the incremental cost utility of docetaxel versus paclitaxel was an estimated Pounds 2431 per quality-adjusted life-year (QALY) in the UK analysis (1994 values) and US$8615 per QALY in the US analysis (1997 values). The French model found that docetaxel cost 700 French francs less for the extra days of QPFS (1993 costs). CONCLUSIONS: Cost-utility analyses, compiled in the absence of direct comparative data, show that docetaxel offers utility gains versus paclitaxel in the treatment of metastatic breast cancer. The incremental cost for these gains is within the accepted range for healthcare interventions.     

Drug Evaluation: Oncologic,Endocrine & Metabolic: Docetaxel (Taxotere®): current status and clinical prospects

New active anticancer agents are found only sporadically, and docetaxel (Taxotere®) serves as an example of how rationalisation can be directly translated into more effective therapy. The heretofore parochial approach to cancer research need not detract from the rational efforts to synthesise active drug analogues, or from designing improved clinical protocols. At the outset, such molecular and pharmacological ‘tinkering’ clued investigators into the structure-activity relationship of anticancer drugs and their more efficacious use. Such is the case for the taxoid, docetaxel. This novel drug tends to be more potent in vitro than paclitaxel (Taxol®) and clinical trials indicate that docetaxel displays activity in what are considered to be poorly responsive tumours (e.g., advanced breast cancer, non-small cell lung cancer, and ovarian carcinoma). Nevertheless, further comparative clinical studies are required to assess the differences between docetaxel and paclitaxel.     

Therapeutic applications of medicinal plants in the treatment of breast cancer: a review of their pharmacology, efficacy and tolerability

Various active compounds (or their semi-synthetic derivatives) derived from medicinal plants have been assessed for their efficacy and tolerability in the treatment of breast cancer. Some of these plant species, including Taxus baccata (paclitaxel, docetaxel), Podophyllum peltatum (etoposide), Camptotheca acuminata (camptothecin) and Vinca rosea (vinblastine, vinorelbine) have well recognized antitumour activity in breast cancer, and have been evaluated in clinical trials. For example, results from recent Phase II/III trials have established docetaxel as the most active single agent in the treatment (first or second-line) of advanced metastatic breast cancer. For other plant species such as Panax ginseng and Allium sativum, antitumour activity has been evaluated in experimental studies using cultured cells and animal models, but the therapeutic potential in patients remains to be determined. Antitumour activity derived from medicinal plants may produce results via a number of mechanisms, including effects on cytoskeletal proteins which play a key role in mitosis (paclitaxel), inhibition of activity of topoisomerase enzymes I (camptothecin) or II (etoposide), stimulation of the immune system (Viscum album), or antiprotease-antioxidant activity. Medicinal plant-derived antineoplastic agents may be used in single agent or in combinational therapies, and have been used in first-line or second-line (including anthracycline-refractory patients) treatment of localized or metastatic breast cancer. Adverse effects resulting from the use of these agents include neutropenia and peripheral neuropathies.     

Epidermal growth factor receptor inhibitor (PD168393) potentiates cytotoxic effects of paclitaxel against androgen-independent prostate cancer cells

Recent data showed that epidermal growth factor receptor (EGFR) inhibitors, such as ZD1839, alone or in combination with chemotherapeutic agents for androgen-independent prostate cancer (AIPC) did not produce promising results in clinical settings. More effective regimens involving novel stronger inhibitor of EGFR and better combinations are needed. The anti-tumor activity of PD168393, an irreversible EGFR inhibitor, with or without chemotherapeutic agents for the treatment of AIPC was investigated in vitro. In results, both the androgen-independent cell lines PC-3 and DU145 expressed higher levels of EGFR than the androgen-dependent MDA PCa 2b and androgen-responsive LNCaP cells by Western blotting. DU145 was much more sensitive to PD168393 and ZD1839 than MDA PCa 2b. PD168393, but not ZD1839, significantly potentiated paclitaxel cytotoxicity against DU145 by MTT assay and median-effect analysis. The combination of PD168393 or ZD1839 with other cytotoxic agents including docetaxel and 5-fluorouracil, however, was either additive or antagonistic. Compared to paclitaxel alone, PD168393 significantly enhanced paclitaxel-induced DNA fragmentation, sub-G1 fraction accumulation, mitochondrial membrane dysfunction, cytochrome C release, caspase-3 activation and eventually apoptosis. These molecular events were accompanied by Bad up-regulation, p53 and p21Waf1/Cip1 induction, ERK1/2 inactivation and inhibition of EGFR phosphorylation in the presence of PD168393. These effects did not involve significant alteration in the Akt1/2 and STAT3 signaling pathway. In conclusion, the combination of paclitaxel and PD168393 produced a profound synergistic growth inhibition of AIPC cells. Combining PD168393 with paclitaxel may have clinical benefits and warrants further investigation.